title: Metabolic activation of aromatic amine mutagens by simultaneous expression of human cytochrome P450 1A2, NADPH-cytochrome P450 reductase, and N-acetyltransferase in Escherichia coli.
abstract: We describe the construction of a new strain of Escherichia coli designed to bioactivate aromatic amines and to detect their mutagenicity with high sensitivity. Strain DJ4309 bears two plasmids, a pACYC184-derived plasmid which expresses Salmonella typhimurium acetyl CoA:arylamine N-acetyltransferase (NAT) and a pBR322-derived plasmid which expresses human cytochrome P450 1A2 and NADPH-cytochrome P450 reductase. The combined actions of these enzymes convert aromatic amines into reactive, mutagenic N-acetoxy esters. The strain also carries a mutated copy of the lacZ gene (on an F' factor) which reverts to the wild-type gene by a -(GpC) frameshift mutation. Strain DJ4309 expresses high levels of NAT and cytochrome P450 1A2 and is very sensitive to mutagenesis induced by representative aromatic amines. Mutagenicity of 2-aminoanthracene in strain DJ4309 is higher than can be obtained by rat liver homogenate 9000g supernatant (S9) activation in the parent strain lacking the P450 expression vector. Strain DJ4309 provides a useful system for detecting mutagenic aromatic amines and for studying their metabolism by human P450 1A2.
authors.affiliation: Guelph-Waterloo Centre for Graduate Work in Chemistry, Department of Chemistry and Biochemistry, University of Guelph, Ontario, Canada. josephy@chembio.uoguelph.ca
authors.forename: P D
authors.initials: PD
authors.lastname: Josephy
authors.forename: D H
authors.initials: DH
authors.lastname: Evans
authors.forename: A
authors.initials: A
authors.lastname: Parikh
authors.forename: F P
authors.initials: FP
authors.lastname: Guengerich
mesh_terms: D001191:Arylamine N-Acetyltransferase / Q000378:metabolism*; D001711:Biotransformation; D019388:Cytochrome P-450 CYP1A2 / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D006841:Hydrocarbons, Aromatic / Q000378:metabolism*; D009153:Mutagens / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase; D010957:Plasmids
chemical_list: D006841:Hydrocarbons, Aromatic; D009153:Mutagens; D019388:Cytochrome P-450 CYP1A2; D009247:NADH, NADPH Oxidoreductases; D009251:NADPH-Ferrihemoprotein Reductase; D001191:Arylamine N-Acetyltransferase